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  • 1
    Electronic Resource
    Electronic Resource
    Microtubules with altered assembly kinetics have a decreased rate of kinesin-based transport (1994)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 27 (1994), S. 79-87 
    Details
    ISSN: 0886-1544
    Keywords: microtubule transport ; microtubules ; 2,5-hexanedione ; glutaraldehyde ; kinesin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Microtubules treated with the γ-diketone 2,5-hexanedione (2,5-HD) have altered assembly behavior characterized by precocious nucleation and rapid elongation. By measuring the rate of microtubule transport, we have examined the potential functional significance of this 2,5-HD-induced microtubule modification. 2,5-HD-treated microtubules were transported at only 70% of the rate of control microtubules in a simple kinesin-based motility assay on glass coverslips using video and computer enhanced differential interference contrast microscopy. Since 2,5-HD is capable of forming both pyrrole adducts and crosslinks with tubulin, the contributions of pyrrole formation and crosslinking to slowed microtubule transport were determined. 3-Acetyl-2,5-hexanedione (AcHD), a pyrrole forming, non-crosslinking congener of 2,5-HD which does not alter microtubule assembly, did not produce slowed microtubule transport as occurs with 2,5-HD. However, glutaraldehyde, a pyrrole-independent crosslinking agent which alters microtubule assembly in the same way as 2,5-HD, slowed microtubule transport. These results indicate that a 2,5-HD-induced microtubule modification, possibly a crosslink-related conformational change, produces both an alteration in the kinetics of assembly and an alteration in the microtubule-motor interaction. © 1994 Wiley-Liss, Inc.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970270109
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  • 2
    Electronic Resource
    Electronic Resource
    Analysis of thymidine kinase gene expression in preimplantation mouse embryos (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 39 (1994), S. 259-267 
    Details
    ISSN: 1040-452X
    Keywords: Thymidine kinase ; Preimplantation mouse embryos ; Embryonic gene expression ; Cell cycle ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Thymidine kinase (TK) activity was examined during the development of preimplantation mouse embryos. TK activity was increased approximately 20-fold from day 2 embryos (2-cell) to day 5 embryos (late blastocyst). TK activity did not change along with the progression into S-phase of the first and the second cell cycles but increased sharply at S-phase of the third cell cycle. Analysis of TK mRNA with a reverse transcription-polymerase chain reaction (RT-PCR) method showed that the level of TK mRNA was low in ovulated eggs and 1-cell embryos and was hardly detectable in day 2 embryo (2-cell), but sharply increased in day 3 embryos (mixture of 5- to 8-cell and morula). The functional role of 5′-flanking sequence of TK gene was also investigated in preimplantation embryos after microinjection with the DNA construct of 5′-flanking sequence of TK (2.4 kb) linked to bacterial lacZ gene (TK2.5lacZ) into the pronucleus of 1-cell and subsequently by histochemical staining with X-gal. β-Galactosidase activity was first detected in day 3 embryos (8-cell), and 30% of embryos were stained with X-gal in day 4 and day 5 embryos, respectively. These results show that an increase in TK activity occurred after 2-cell stage, and this increase was primarily due to the embryonic activation of TK gene expression. Also, it appears that the 5′-flanking sequence of TK may directly regulate the TK gene expression at the transcriptional level during preimplantation murine development. © 1994 Wiley-Liss, Inc.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/mrd.1080390303
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  • 3
    Electronic Resource
    Electronic Resource
    Structural and functional changes in salivary glands during aging (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Microscopy Research and Technique 28 (1994), S. 243-253 
    Details
    ISSN: 1059-910X
    Keywords: Senescence ; Lipofuscin ; Secretion and content of saliva ; Protein synthesis ; Amylase mRNA ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Natural Sciences in General
    Notes: Various salivary glands in senescent humans and other animals have been examined extensively to characterize the structural and functional changes that occur during aging. Although a wide range of different structural changes, involving both the parenchymal and stromal tissues, have been described, it is unclear how any of these changes affects the function of the salivary glands. One major change in structure is the reduction in the volume of acini with a concomitant increase in the ductal volume. Despite this loss of functional acini, the salivary output and the contents seem to be unaltered, or minimally altered, due to aging. One consistent change observed in many salivary glands of aged animals is the decline in the rate of synthesis of proteins and their messenger RNA (mRNA). However, the salivary acinar cells from aged animals can synthesize secretory proteins at an elevated rate just as effectively as those from their younger counterparts in response to external stimuli, which are known to enhance the rate of protein synthesis. Thus, it appears that the salivary acinar cells, which remain structurally intact during aging, seem to retain their functional efficiency. Furthermore, these acinar cells, although reduced in number, are sufficient in quantity to carry out most of the salivary gland functions. © 1994 Wiley-Liss, Inc.This article is a US Government work and, as such, is in the public domain in the United States of America.
    Additional Material: 14 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jemt.1070280308
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  • 4
    Electronic Resource
    Electronic Resource
    Inhibition of angiogenesis by tissue inhibitor of metalloproteinase (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 160 (1994), S. 194-202 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Matrix proteases play a critical role in cell invasion and migration, including the process of angiogenesis. The ability of specific factors to induce angiogenic responses correlates with their stimulation of matrix protease synthesis and release. Using an in vivo angiogenesis assay, the endothelial cell response to known angiogenic factors, basic fibroblast growth factor (bfGF) and adipocyte conditioned medium, was blocked by an inhibitor of matrix metalloproteinase activity, TIMP-1. The TIMP effect was mediated, at least in part, through the inhibition of endothelial cell migration, as determined by the ability of TIMP to block chemotaxis in a Boyden chamber assay. These results indicate that the inhibition of migration is a direct effect on the endothelial cells and does not require accessory cells. An additional observation was that the RNA levels for TIMP were significantly reduced in differentiated adipocytes, compared to undifferentiated F442A controls. Therefore, the acquisition of an angiogenic phenotype may involve not only the induction of positive factors, but also the suppression of angiogenesis inhibitors. © 1994 Wiley-Liss, Inc.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041600122
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  • 5
    Electronic Resource
    Electronic Resource
    Mouse fibroblasts defective in thrombin mitogenesis possess functional proteolytically activated receptor for thrombin: Requirement for a second signaling pathway (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 160 (1994), S. 573-584 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Thrombin mitogenesis in fibroblasts requires two distinguishable subsets of signals; one generated by proteolytic cleavage, the other by high-affinity cell surface binding. Characterizing two closely related mouse embryo (ME) cell lines with high numbers of thrombin binding sites, we found that one line, B11-A, responds mitogenically to thrombin, epidermal growth factor (EGF), and serum, whereas the B11-B cell line is responsive to EGF and serum, but not to thrombin. The B11-B defect responsible for loss of thrombin responsiveness is not due to differences in the number of high-affinity binding sites, the affinity of thrombin binding to these sites, or to differences in cell surface expression of proteolytically activated receptors for thrombin (PART). The defect is also not associated with an inability of thrombin to activate PART since thrombin stimulates the cleavage-dependent induction of the proto-oncogene c-fos in both B11-A and B11-B cells. Various combinations of thrombin, synthetic thrombin receptor peptide, TRP-14 (SFFLRNPGENTFEL), platelet-derived growth factor (PDGF), and phorbol 12-myristate 13-acetate (PMA) were used to better define the defect in thrombin-mediated mitogenesis in B11-B cells. Direct activation of protein kinase C with PMA in combination with thrombin did not overcome B11-B nonresponsiveness. However, mitogenic responsiveness was regained in B11-B cells by simultaneous addition of PDGF and either thrombin or TRP-14. Therefore, the B11-B defect may involve a set of signals initiated by nonproteolytic thrombin interactions distinct from those initiated by PART, but related to the downstream signals initiated by the tyrosine kinase-associated growth factors, EGF and PDGF. © 1994 Wiley-Liss, Inc.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041600321
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  • 6
    Electronic Resource
    Electronic Resource
    Stable differentiation of a human colon adenocarcinoma cell line by sodium butyrate is associated with multidrug resistance (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 160 (1994), S. 213-226 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Colorectal cancers are often composed of cell types representing various differentiated cell lineages, however little is known concerning the relationship of differentiation and drug resistance in these cancers. The present study was performed to develop and characterize a stable, differentiated clone of the human colon cancer cell line LS174T and to characterize the drug resistance of this cell line in relation to its undifferentiated parental cell line. LS174T cell line was treated with the differentiating agent sodium butyrate (0.5 mM) for 30 days, then recultured in standard medium. Foci of flat-appearing cells appeared and were isolated using cloning rings, and subcloned. One subclone was designated LS174T-D. The LS174T-D clone maintains a stable, differentiated phenotype in standard culture conditions in the absence of sodium butyrate. It is characterized by the formation of a polarized monolayer with dome formation and the presence of prominent apical microvilli and tight junctions. This cell line demonstrated reduced growth in soft agar and nude mice compared with the parental cell line. LS174T-D cells expressed immunoreactive intestinal mucin antigens and brush border enzymes dipeptidyl aminopeptidase (DAP)-IV and aminopeptidase. The activities of DAP-IV and aminopeptidase were increased 5.6-fold and 3.4-fold, respectively, in LS174T-D compared with parental cells. Proliferation assays demonstrated that, compared with the parental cell line, LS174T-D cells were more resistant to doxorubicin (93-fold), cisplatin (23-fold), 5-fluorouracil (12-fold), 5-fluorodeoxyuridine (31-fold), and methotrexate (12.5-fold). Intracellular uptake of (3H)-5-fluorodeoxyuridine did not differ significantly in the differentiated and undifferentiated cell lines. Levels of mdr-1 p-glycoprotein measured by Western blot and RNA Northern blot assays were also similarly low in both cell lines. However, total glutathione content and glutathione-S-transferase activities were increased in LS174T-D cells by sixfold and threefold, respectively, compared with parental cells. Depletion of glutathione by pretreatment with DL-buthionine sulfoximine reversed LS174T-D resistance to cisplatin. Long-term treatment with sodium butyrate induces or selects for colon cancer cells with features of enterocytic differentiation. This stably differentiated cell line is associated with glutathione-mediated multidrug resistance, and provides a model for further studies of differentiation in normal and cancerous colon. © 1994 Wiley-Liss, Inc.This article is a US Government work and, as such, is in the public domain in the United States of America
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041600202
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  • 7
    Electronic Resource
    Electronic Resource
    Transforming growth factor-β regulation of retinoblastoma gene product and E2F transcription factor during cell cycle progression in mouse fibroblasts (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 160 (1994), S. 1-9 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The mechanism by which transforming growth factor beta (TGFβ) exerts growth stimulatory effects was examined in C3H/10T1/2 mouse fibroblasts by study of cell cycle regulation of the retinoblastoma gene product (p110Rb) and transcriptional regulation of the p110Rb-associated transcription factor, E2F. Northern blotting analysis shows that TGFβ and/or epidermal growth factor (EGF) stimulate by three to sixfold the level of Rb mRNA which is also reflected by the increased levels of p110Rb. p110Rb becomes phosphorylated in mid-G1 and further phosphorylated at the G1/S transition. Hyperphosphorylation of p110Rb by TGFβ can be observed when cells are in S phase. TGFβ stimulates by three to fourfold the activity of cdk2 kinase consistent with the observed phosphorylation of p110Rb and also with the possibilit that the kinase is involved in phosphorylating p110Rb close to the G1/S transition. Thus, TGFβ as a growth stimulator induces, as does EGF, the phosphorylation of p110Rb during cell cycle progression. Transient transfection of E2F promoter constructs was used to analyze the effect of TGFβ on the modulation of E2F-mediated transcription. The data revealed that TGFβ can stimulate wild-type adenoviral E2 promoter activity by 12-fold. Taken together, TGFβ-induced phosphorylation of p110Rb in mouse fibroblasts appears to exert a positive regulatory function upon genes that have a pivotal role in the G1/S transition of the cell cycle. © 1994 Wiley-Liss, Inc.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041600102
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  • 8
    Electronic Resource
    Electronic Resource
    II. Yeast sequencing reports. The sequence of a 32 420 bp segment located on the right arm of chromosome II from Saccharomyces cerevisiae (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 10 (1994), S. S47 
    Details
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; yeast ; chromosome II ; RIF1 ; DPB3 ; MRP-L27 ; SNF5 ; SEC61-homolog ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The sequence of a 32 420 bp segment of Saccharomyces cerevisiae chromosome II has been deduced. The sequence data revealed 19 potential new genes covering 83·5% of the sequence. Four genes had already been cloned and sequenced: part of RIF1, DPB3, MRP-L27 and SNF5. Besides these four genes, 15 open reading frames (ORFs) of at least 100 amino acids encoding potential new genes were identified. Two of these ORFs are overlapping and a third is located within another ORF.The putative gene product of ORF YBR2039 was homologous to the group of uncoupling proteins involved in the mitochondrial energy transfer system. We propose a remapping of the MRP-L27 gene encoding the mitoribosomal protein YmL27 as it previously has been mapped on chromosome X. The ORF YBR2020 has a strong homology with a 31·9% identity in a 473 amino acid region to the yeast gene SEC61, suggesting that YBR2020 is a new gene encoding a protein involved in translocation of proteins in the yeast cell. Six of the potential genes do not exhibit any significant homology to previously sequenced genes as predicted in the Fast A analysis. The sequence has been deposited in the EMBL data library under Accession Number X76053.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/yea.320100007
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